A Validation Week

Sometimes I think that having a holiday is what you need to guilt you back into working. Last weekend I went home (together with 3 friends from Ireland) for an extended weekend. I really enjoyed showing them the sights of Malta, and also meeting my sister who came over too. However once I got back into the lab I knew that I needed to get down to business. Since the Mass Spec is still not working (still waiting for the parts to arrive), I decided to make use of the HPLC part of the LC-MS instrument. Usually you are not allowed to make use of this front end portion of the instrument on its own. However since the MS part was not working I got permission to use it. Therefore this gave me time to work on my method’s transferability to the actual instrument I will be (hopefully) using.

Before my trip I had ascertained that the method transfers quite well, and I was pleased with the preliminary results. However I knew that as with any method this needs to be validated. Therefore on my return I started working on this section of the process. I have now finished the validation of the HPLC method and I am quite pleased with the overall results. I did have to remove an analyte from the 11 I had started with, since it did not validate in any test. But I am not too concerned about that. After all that is why I started with 11. My supervisors had initially told me not to go for more than 5 analytes, but I had argued that it is better to start with more, and then eliminate later on, depending on the results.

I certainly hope that the parts arrive soon, and that the instrument is fixed very soon. In the meantime, I have figured out what I could do to ‘fill the time’. The parts have been said to be delivered either yesterday (so no) or early next week. I desperately hope that these parts are the actual ones required and that once it is fixed it keeps working. Another girl who is finishing her PhD will probably get onto the instrument first. But as long as it is working there is hope.

Getting back into the swing of things

After my Easter holiday, I was finding it quite hard to get back to work, so that last week I mainly did paper work and no experimental work as such. I did not feel as productive, but I did manage to finish all the documentation (reports, poster, plans) for next week’s ATWARM meeting in Germany.

On a positive note, I had a meeting with the supervisor who had not yet seen my literature review last Wednesday, and he seemed pleased with the way it looks. He suggested some further changes which I also finalised last week and I resent it to him as requested and am now awaiting further comments. Hopefully he approves with the changes so that I could resend it to all the supervisors and can get the ball rolling.

Last week I also got the new HPLC column I ordered just before going on holiday. I will get to working on the HPLC tomorrow to see how this works out using the previous conditions. In theory they are identical columns, just that this one is new and therefore should be ‘better’. But I am learning to expect the unexpected, so will wait and see.

Hopefully this week is more productive, and I get some actual lab work done, since I will be away most of next week for the meeting in Germany.

Research Progresses

After the ENVIRON conference I got back into my HPLC method development and literature review work. Regarding my literature review my supervisors and I have decided on a target journal for submission, and I have looked at the guide for authors and formatted my document to their requirements e.g. reference style, sections etc. At least Refworks, which I use as a reference manager, had the journal style inputted, and it corresponded to the journal’s requirements, and therefore it was easy enough to achieve. I have now sent my review to a supervisor who has not yet seen it and had requested to see it after I carry out the changes of the three supervisors who had seen the previous draft.

HPLC method development has also progressed well. I have now separated 10 of the analytes (out of 11), with good peak separation and got a good calibration curve. For two of the analytes (i.e. analytes 10 and 11) no peak separation occurs. However they absorb at different wavelengths and therefore I can use wavelength separation for them. Unfortunately the HPLC I am currently using doesn’t have a DAD. However the LC-MS detector has a DAD and therefore I could save time from having to do all analyses twice. I have discussed the HPLC method with one of my supervisors who knows the area most, and he gave me some suggestions re some ghost peaks (small peaks coming out even in the blank), and I will try them out in the coming weeks. Once that is finished, I hope that the LC-MS would have been fixed and I can get onto it (it has been broken down since before the 10th March, so we will see).

Today I am off for an Easter trip around Ireland with the rest of the family who are visiting from all over Europe. I can’t wait to get a break and be back and work!

Some progress is made

Finally, we managed to get the HPLC up and running again (basically I managed to convince the technician to change the tubing to a different type), so I had my first run yesterday, which kept going till this morning. The results are still not totally there, but the R squared values are all higher than 0.9, and those that are low are due to known reasons. So that is a good thing. I should be getting back on the equipment next week for at least 2 days, so that is something to look forward to, and hopefully I can get some good results and progress to the next stage.

Today I also received the last of the comments from my supervisors about my literature review (received from 3 out of 5, which is the most I can expect at the moment: one is out on long-term sick leave and the other said he will see the next draft, which is better after all, because I would not have to join all comments together). That is on the agenda for next week too. Overall the comments were quite manageable. They should not involve too much work, and hopefully I can get through them in a week or so, since most involve removing certain sections, or particular quick to do word changes etc.

However, the supervisors suggested that I look at publishing my literature review, which is what is worrying me the most. They have asked me to suggest journals in which to publish my literature review, and I have no idea where to start looking. I have never published papers before in any journal, so I am at a bit of a loss. They talked about impact factors for the journals, and they said it should not be too low, but I do not want to aim too high. I also know that it is difficult to get literature reviews published, unless it is invited (which this certainly is not). We’ll see how things go.

An unproductive week

Another PhD week is over, and it can certainly be labelled as unproductive. I submitted the second draft of my literature review to my supervisors last Monday (1st time all except one are seeing it) and that is practically the extent of my work. The problem is that I have been planning on working on the HPLC method development, but I am having big problems with that. Some tubing has been changed in the past couple of weeks, and therefore big problems with pressure are ensuing, resulting in the tubing popping off.

I hope next week I can be a bit more productive and get my HPLC method working. If I can get back to how I was a couple of weeks ago on my method, it should only take a couple of days (2 or 3) to get everything I want done done. But Oh well.

Wish me luck for the coming week.

More HPLC Woes

My HPLC woes continue. I have finally gotten onto the HPLC to do some work, but have not been getting any peaks whatsoever.  I tried a couple of different approaches, but to no avail. Being sick over the past few days certainly does not do anything to help me think or to boost my morale. After carrying out a couple of runs this morning, I decided to take the rest of the day off and have a good rest. I turned over the HPLC to the next person who needs to use it, and hopefully can get access to it, and some work done tomorrow.

On a positive note, I managed to finish my second draft of my literature review yesterday and sent it to all my supervisors for comments and advice. I had sent the first draft to just one supervisor, and had gotten some constructive comments on how to improve it. I will await comments on this draft hopefully in the coming weeks.

Lab Meeting

On Thursday I managed to squeeze in a lab meeting with two of my supervisors on Thursday between the two lab demonstration sessions. Rather than a lab meeting where all of us are present, it was an individual meeting where only myself, the two supervisors (the other is away sick) and the lab post-doc was present. I had not had such a lab meeting in more than a month, so it is good to give a presentation on progress.

First up for discussion was the literature review. One of the supervisors had seen it and all that I had done was work on the changes she suggested. However the other knew that I was working on it, but not on the progress. The changes are by no means completely done, but hopefully I can polish it up somewhat in the coming weeks. I am pleased that there were not many changes suggested by my supervisor, though there are a couple of areas I feel I need to improve.

I then discussed my method development. I have now nearly determined the method for the first few analytes. On Thursday, after the meeting I also received the next few analytes, so what I am doing today is trying to include them in the same method, to reduce sample preparation and analysis time. It was good to present some actual findings in the meeting, and got some good suggestions from one of my supervisors, who is a chemist and knows a whole lot about everything.

As with all of my meetings, it was a short and sweet meeting. I hope I manage to have some good results for the next meeting, whenever that will be. Hopefully I will have nearly finalized my HPLC method, so I can go on to SPE and LC-MS method development. I have some analysis currently running, so hopefully by the end of the weekend I will know a bit better about how much more work is needed. Wish me luck

Another Week is Over

The past week has been a hectic week PhD wise. I had a conference on Monday and Tuesday, a meeting with my supervisor on Wednesday and lab demonstrations on Thursday. In addition I have been working to develop my HPLC Method. I have spoken about the conference and lab demonstrations previously. I have also touched upon the HPLC method development.

The method development has been progressing slowly but steadily. If things keep on progressing the way they are I would be indeed quite happy. I tried out some things suggested by a girl I met at the conference last Tuesday, and things have improved. I did some more runs yesterday (yes I know it was a Saturday) and got the results this morning (yes it is a Sunday), and I am quite pleased with the way things are progressing, though 1 peak is still missing/it peeks out a bit from behind another, so that it on my agenda for tomorrow. The R squared values for the calibration curve I did are all greater than 0.99, so not too bad, though there is room for improvement. Once I (hopefully) separate my current 6 peaks, I will then need to add in another 5 compounds (ordered this week ), and then start working on joining the HPLC method to the sample preparation technique and the Mass Spec analysis technique. So still a way to go to be able to start collecting samples for analysis. But we will get there.

I also had a meeting with my supervisor on Wednesday. We generally have lab meetings every Thursday, but since I am demonstrating all day Thursday, I will be having meetings with my supervisor every now and then depending on need, until I finish demonstrating. During the meeting we discussed a couple of things I had been meaning to talk to her about, in particular things which do not lend themselves well to discussing during a lab meeting.

First up was the thing I was dreading most. I had given her my literature review a couple of weeks ago (just before she went on holiday, and she read it during her holiday). I was quite pleased with the feedback however. She said it was quite good for a first try (that is what we aim for after all), and she gave me a couple of changes to make. I have now carried out all the minor changes, and will start focusing on the sections which involve searching for more data and so on. The main issue I need to do is to summarize  everything  at the end, rather than simply saying what other people said, without giving any opinions, and therefore this leaves everything hanging a bit. She also noticed my love of the words therefore, nevertheless etc, and was told to cut down on those. The last part of the meeting consisted of me discussing some points I wanted to talk to her about, such as ordering of the new chemicals, how my progress is getting along, the conference details, and lastly, but quite importantly for me, we discussed time off. I now have approval for my days off till the end of August. It feels good to know roughly when I can get to go home etc etc.

Overall, a busy but pleasant week. I could do with it being slightly less hectic, but rather a week like this one, to a slow week, or one when nothing goes well.

HPLC Method Development

I have been working on trying to develop an HPLC method since January. However things have not been really all that successful (I have mainly just been cleaning the column, meaning that I have achieved nothing basically). However last week I decided to change tracks and try something else, where I develop my own method from scratch, rather than using a ready made method. And it seems to work. I tried to separate and detect 4 compounds and managed successfully, although I need more work to separate 2 compounds which have similar retention times. However that should not be such a big problem.

I got my first results yesterday, and I must say I was pleased. I still need to add in a couple of other (around 10 more) compounds hopefully, but I am optimistic. I also got good calibration curves for both the single injections and the mixed injections (although I also got some horrible ones, which I am however attributing to incorrect sample preparation, since it is generally one point which is off rather than a whole set.

I will next get time on the HPLC instrument next Wednesday, since I will be at a conference on Monday and Tuesday, and seeing as we are now 4 people using the same instrument, we need to share time efficiently. However I am now a bit more optimistic about the method I am using, so I am happy enough for now. I wished to have been at this point a couple of weeks ago, but Oh well, here we are now.

Graduate Training

Last week saw me attending my first module from the Graduate Training programme at DCU: Advance Analytical Techniques. The course consisted of a number of lab sessions where all students get to perform 4 experiments of their choice. Students (mainly in chemistry) from all the Dublin Universities (DCU, UCD, Trinity, NUIM) were participating as part of the Dublin Region Higher Education Alliance. The week was quite hectic, having to perform an 6 hour experiment each day, followed by a lab report to be written.

Overall, I must say the week was quite fruitful. I got back into the lab routine, such that you remember (or are reminded of) all the small nitty gritty things you have forgotten and all the other things you have never heard about. It was also good to meet up with all the people from the various Universities, and find out a bit about what they are doing, and also discuss some differences between the research environment at different locations. In addition, our demonstrator in charge (Dr Blanaid White) was really knowledgeable in all the experiments, and we also got the expertise of other individuals from the School of Chemistry, to give us further information about a number of experiments about which they have lots of experience.

The experiments I performed were about GC-MS, HPLC, Cyclic Voltammetry and Ion Chromatography. From each experiment I learnt a lot, but I was most excited about the HPLC experiment, since I will be working a lot with HPLC during my PhD, and this session helped me understand more about how the different parameters function and the basics about how to develop a method.

The lab reports are now written and I will be submitting everything on Monday. Let us hope I did well, as it would mean that the past week has indeed been fruitful. I wish that DCU has even more graduate training elements from which to choose from, as this has certainly been useful. I will be doing some more in the coming months, and I also hope they will be as fruitful as this.